Dipotassium glycyrrhizate prevents oral dysbiosis caused by Porphyromonas gingivalis in an in vitro saliva-derived polymicrobial biofilm model.

OBJECTIVES: Oral microbiome dysbiosis prevention is important to avoid the onset and progression of periodontal disease. Dipotassium glycyrrhizate (GK2) is a licorice root extract with anti-inflammatory effects, and its associated mechanisms have been well-reported. However, their effects on the oral microbiome have not been investigated. This study aimed to elucidate the effects of GK2 on the oral microbiome using an in vitro polymicrobial biofilm model. METHODS: An in vitro saliva-derived polymicrobial biofilm model was used to evaluate the effects of GK2 on the oral microbiome. One-week anaerobic culture was performed, in which GK2 was added to the medium. Subsequently, microbiome analysis was performed based on the V1-V2 region of the 16 S rRNA gene, and pathogenicity indices were assessed. We investigated the effects of GK2 on various bacterial monocultures by evaluating its inhibitory effects on cell growth, based on culture turbidity. RESULTS: GK2 treatment altered the microbiome structure and decreased the relative abundance of periodontal pathogenic bacteria, including Porphyromonas. Moreover, GK2 treatment reduced the DPP4 activity -a pathogenicity index of periodontal disease. Specifically, GK2 exhibited selective antibacterial activity against periodontal pathogenic bacteria. CONCLUSIONS: These findings suggest that GK2 has a selective antibacterial effect against periodontal pathogenic bacteria; thus, preventing oral microbiome dysbiosis. Therefore, GK2 is expected to contribute to periodontal disease prevention by modulating the oral microbiome toward a state with low inflammatory potential, thereby utilizing its anti-inflammatory properties on the host.

An open-label, investigator-initiated, single-center, prospective, pilot clinical study to evaluate the efficacy of a skin whitening serum applied twice daily combined with a spot-preventing SPF50+ sunscreen in healthy female subjects with melasma hyperpigmentation.

BACKGROUND: Melasma is a common skin disorder characterized by alterations in normal skin pigmentation. The objective was to evaluate the efficacy and safety of a skin whitening serum containing niacinamide, hydroxyphenoxy propionic acid, dipotassium glycyrrhizate, glycolic acid, and 4-n-butylresorcinol applied twice daily combined with a spot-preventing SPF50+ sunscreen for treatment of melasma. METHODS: Twelve healthy Caucasian women with melasma (Fitzpatrick skin types II-IV) were enrolled in this pilot clinical study. Efficacy evaluations were performed at baseline and weeks 4, 8, and 12 of treatment and included clinical and instrumental assessments. RESULTS: All endpoints for melasma hyperpigmentation showed a statistically significant improvement from baseline to the end of the study. There was only one dropout. No signs of irritation or discomfort were observed at baseline, w4, w8, or w12. An overall improvement in melasma was observed both clinically and on reflectance confocal microscopy (RCM). CONCLUSION: This topical skin whitening serum had favorable outcomes for the treatment of melasma hyperpigmentation in adult women, as demonstrated on investigator and instrumental assessments. The results of this pilot study need to be confirmed in randomized, controlled studies with a larger sample size.

Restoration of calcium-induced differentiation potential and tight junction formation in HaCaT keratinocytes by functional attenuation of overexpressed high mobility group box-1 protein.

HaCaT cells have been widely used as undifferentiated epidermal keratinocytes, since these non-tumorigenic cells can be readily maintained in conventional medium and partly retain epidermal differentiation potential upon stimulation with high concentration of calcium. In contrast to primary epidermal keratinocytes, however, these cells never form tight junction (TJ), a specific structure in highly differentiated keratinocytes, solely by the differentiation stimulation. Here, we show that HaCaT cells secrete a considerable amount of high mobility group box-1 protein (HMGB1), one of major inflammatory mediator, which appeared to be responsible, at least in part, for such aberrant differentiation response. So far, inhibition of c-Jun N-terminal kinase (JNK) in high calcium medium has been supposed to be the only way to induce TJ formations in HaCaT cells; however, SP600125, a potent inhibitor of JNK showed cytostatic effects and clearly attenuated epidermal differentiation and stratification. In contrast, dipotassium glycyrrhizate (GK2), a soluble analogue of HMGB1-blocker Glycyrrhizin, down-regulated interferon-ß, a typical inflammatory cytokine induced by secreted HMGB1, and accelerated differentiation responses to the calcium treatment in these cells. In addition, GK2-treatmenrt resulted in the formation of double cell layers in cultured HaCaT cells, where the stratified upper cells transiently accumulated TJ proteins at the cell-cell contact sites. These results highlight the importance of attenuation of secreted HMGB1-signals in cultured HaCaT cells for studies of functional keratinocytes.

Dipotassium Glycyrrhizate Improves Intestinal Mucosal Healing by Modulating Extracellular Matrix Remodeling Genes and Restoring Epithelial Barrier Functions.

Gut mucosal healing (MH) is considered a key therapeutic target and prognostic parameter in the management of inflammatory bowel disease (IBD). The dipotassium glycyrrhizate (DPG), a salt of the glycoconjugated triterpene glycyrrhizin, has been shown to inhibit the High Mobility Group Box 1 (HMGB1) protein, an allarmin strongly implicated in the pathogenesis of most inflammatory and auto-immune disorders. Here we discuss new insights on how DPG acts on MH comparing the acute phase and the recovery phase from experimental colitis in mice. We found that DPG strongly accelerates MH by differently regulating pro-inflammatory (CXCL1, CXCL3, CXCL5, PTGS2, IL-1ß, IL-6, CCL12, CCL7) and wound healing (COL3A1, MMP9, VTN, PLAUR, SERPINE, CSF3, FGF2, FGF7, PLAT, TIMP1) genes as observed only during the recovery phase of colitis. Relevant issue is the identification of extracellular matrix (ECM) remodeling genes, VTN, and PLAUR, as crucial genes to achieve MH during DPG treatment. Furthermore, a noticeable recovery of intestinal epithelial barrier structural organization, wound repair ability, and functionality is observed in two human colorectal adenocarcinoma cell lines exposed to DPG during inflammation. Thus, our study identifies DPG as a potent tool for controlling intestinal inflammation and improving MH.

Dipotassium glycyrrhizate via HMGB1 or AMPK signaling suppresses oxidative stress during intestinal inflammation.

AIMS: Oxidative stress and inflammation are always associated. Appropriate management of oxidative mediators may represent a therapeutic strategy to reduce inflammation, and use of antioxidant can be protective against inflammatory diseases. Glycyrrhizin (GL) plays an anti-inflammatory and antioxidant effect by inhibiting high mobility group box 1 (HMGB1) or 11-ß-hydroxysteroid dehydrogenase type II (11ßHSD2) enzyme. In this study, the potential role of dipotassium glycyrrhizate (DPG), a salt of GL, to reduce oxidative stress in intestinal inflammatory condition was investigated in vivo and the mechanism of action of DPG was studied in vitro. RESULTS: In a colitis mouse model DPG affected oxidative stress reducing iNOS and COX-2 expression, as well as NO and PGE2 levels. By means of LPS-stimulated macrophages we found that DPG inhibited the expression of pro-inflammatory cytokines and reduced iNOS and COX-2 expression in a time dependent manner, through two different ways of signal. DPG reduced, at a later time, both iNOS and COX-2, through a mechanism HMGB1-dependent, and at an earlier time only COX-2, through a mechanism AMP-activated kinase (AMPK)-phosphorylation-mediated. CONCLUSION: DPG has a protective effect on colitis and inflammation through the inhibition of oxidative stress. This study clarifies the two-ways mechanism by which DPG inhibits iNOS and COX-2 during inflammation and demonstrates for the first time that AMPK is a target of DPG. Uncovering this mechanism is significant to clarify the relationship between energy homeostasis and anti-oxidative responses and suggests that DPG could play a relevant role in the development of new therapy against inflammatory diseases associated to oxidative stress.